| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
B in Allergen-Induced T Cell Chemotaxis by IL-16 and RANTES1
2,*
*
Division of Respiratory Cell and Molecular Biology, University Medicine, Southampton University General Hospital, Southampton, United Kingdom; and
Pulmonary Center, Boston University School of Medicine, Boston, MA 02118
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
B
activation. These results indicate that allergen stimulates cells
within the bronchial mucosa to increase IL-16 and RANTES release, both
of which contribute to T cell accumulation in asthmatic airways. The
allergen-induced chemotactic activity is dependent on cell activation
via CD28 and involves, at least partly, NF-B. |
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
A range of cytokines are known to induce lymphocyte chemotaxis,
including IL-16; RANTES; macrophage-inflammatory protein (MIP)-1
and
-1ß; monocyte chemoattractant protein-1, -2, and -3; IL-1; IL-8;
lymphotactin; and IL-15 (6, 7). A number of these are
present in increased quantities in BAL and biopsies of asthmatic
subjects from both allergen-challenged and unchallenged airways.
However, a functional link between the detection of these cytokines and
their role in T cell chemotaxis is far from being fully established. In
our previous study of T cell chemotaxis in asthma, IL-16 and MIP-1,
but not IL-8 and RANTES, were shown to be important for the early
accumulation of T cells in the airways 6 h after local
allergen challenge via the fiberoptic bronchoscope
(8).
The aim of the present study was to extend the investigation to events that occur during 24 h of stimulation with allergen and to begin elucidating the mechanisms that regulate the generation of T cell chemotactic activity. To that effect, we have developed a bronchial explant model that enables mucosal tissue, obtained by bronchoscopic biopsy, to be challenged with allergen ex vivo and the supernatants to be analyzed for cytokine levels and T cell chemotactic activity. We have hypothesized that although preformed IL-16, stored and released by CD8+ T cells (9) or epithelial cells (10), may be important in initiating the early influx of T cells seen 6 h postchallenge, stimulation by allergen for 24 h may reveal an additional contribution of RANTES. By way of its preferential effects on CCR3 chemokine receptors, RANTES may be of particular relevance to atopic asthma in causing selective recruitment of Th2-type T cells and eosinophils.
We have also investigated the effect of dexamethasone on the expression
of RANTES and IL-16 in view of the established therapeutic effects of
corticosteroids in asthma. Although corticosteroids are known to
inhibit the expression of RANTES in bronchial epithelial cells
(11, 12) and airway smooth muscle cells (13)
through a process involving interaction of glucocorticoid receptors
with the transcription factor NF-B and AP-1 (14),
little is known about their effect on IL-16.
We have exploited the explant model for its unique suitability to test
novel concepts of immunoregulation using substances that have not been
approved for human use. We have thus used the proteasome inhibitor
Cbz-Ile-Glu(OtBu)-Ala-leucinal (PSI) (15) to seek evidence
for a role for NF-B in regulating the generation of T cell
chemotactic activity. PSI is an inhibitor of the chymotrypsin-like
subunit of the 20S multicatalytic proteinase of the 26S proteasome
which degrades IB, the cytoplasmic protein that binds NF-B, and
prevents its transport to the nucleus and binding to the promoter
region of NF-B-dependent cytokine genes. By inhibiting the
proteolytic degradation of IB by the proteasome, PSI prevents the
translocation of NF-B to the nucleus and consequently the activation
of NF-B-dependent cytokine genes.
Finally, we have hypothesized that the generation of chemotactic
factors for T cells in the bronchial mucosa requires cell-cell
interaction involving costimulatory molecules. In OVA-sensitized mice,
a central role has been identified for an interaction between CD28,
expressed on T cells, and B7, expressed on APC, for both the
sensitization and secondary Th2-type immune responses to allergen to
occur (16). To test the hypothesis that this interaction
is also important in human asthma, we have used CTLA-4-Ig, a fusion
protein comprised of the Fc fragment of IgG and the extracellular
domain of CTLA-4, which is normally expressed on activated T cells of
both CD4+ and CD8+ subsets
(17). The CTLA-4-Ig fusion protein has been successfully
used to block T cell activation dependent on B7-1 (CD80) and B7-2
(CD86) costimulatory molecules (18). We have recently used
CTLA-4-Ig in our explant model to block allergen-induced production of
IL-5 and IL-13 (19) and have hypothesized that this was
due to an inhibitory effect on T cells. However, in mice, CD28 has also
been shown to be involved in mast cell activation leading to tyrosine
phosphorylation, up-regulation of c-jun and generation of
TNF- and IL-13 (20, 21), but it is not known whether
CD28 is expressed on human mast cells.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Serum-free medium, AIM V for culture of bronchial biopsies, RPMI 1640 for T cell chemotaxis, heat-inactivated FCS, HEPES, L-glutamine, sodium pyruvate, penicillin, streptomycin, Fungizone, and PBS without calcium and magnesium (PBS) were all purchased from Life Technologies (Uxbridge, U.K.). BSA was from Sigma (Poole, U.K.). House dust mite (Der p) allergen extract, containing all the major allergen groups, was from ALK (Horsholm, Denmark). Lymphoprep was from NYCOMED (Oslo, Norway). mAbs for panning (anti-CD11b, -CD14, -CD16, and -CD19) were purchased from Serotec (Oxford, U.K.) and used in conjunction with rabbit anti-mouse Igs (Dako, High Wycombe, U.K.). ELISA kits for the measurement of IL-16 and RANTES were from Lifescreen (Watford, U.K.). Neutralizing polyclonal Abs for RANTES were from R&D Systems (Abingdon, U.K.). Neutralizing anti-IL-16 mAbs were from clone 14.1 used in our previous studies (8). The proteasome inhibitor PSI was purchased from Bachem (Saffron Walden, U.K.), and water-soluble dexamethasone was from Sigma. The fusion protein CTLA-4-Ig was a gift from Dr. Jonathan Ellis (Glaxo Wellcome, Stevenage, U.K.).
Subjects
Seventeen atopic asthmatic subjects (4 females) aged between 21 to 42 years and 12 nonatopic nonasthmatic (control subjects) aged between 23 and 39 years participated in the study. All the asthmatic and none of the control subjects were atopic, as assessed by the presence of positive skin prick tests to one of a panel of common aeroallergens (ALK) and were sensitized to house dust mite (Der p). All had mild asthma with forced expiratory volume in 1 s >70% of predicted. They were treated with the ß2-agonist albuterol as required to relieve asthma symptoms, and none had been on inhaled corticosteroids for at least 2 wk before bronchoscopy.
The study was approved by the Southampton University and Hospitals Ethics Committee, and volunteers gave their written informed consent.
Bronchoscopy and culture of bronchial explants
Bronchoscopy with endobronchial biopsy was conducted according to the National Heart, Lung, and Blood Institute guidelines as previously reported (22). On average six bronchial biopsies were taken of subcarinae, washed twice in AIM V medium, and weighed before culture. One to two biopsies, weighing between 2 and 15 mg, were cultured per well in a final volume of 500 µl of either AIM V medium alone (control culture) or medium to which Der p allergen extract was added at 2000 U/ml. After 24 h of culture, supernatants were harvested, centrifuged at 600 x g for 10 min at 4°C, and stored at -70°C for chemotaxis experiments and cytokine measurements.
Additional cultures were set up using bronchial biopsies from atopic asthmatic subjects to which PSI, CTLA-4-Ig fusion protein, or dexamethasone were added to the culture medium in which tissue was stimulated with allergen. PSI was dissolved in DMSO at 50 mM and diluted in AIM V medium to give a final concentration of 5 µM. Bronchial biopsies were preincubated with 5 µM PSI for 1 h before stimulation with Der p. CTLA-4-Ig was prepared in PBS at 1 mg/ml and diluted with AIM V medium to 25 µg/ml before adding to culture at the same time as the allergen. Water-soluble dexamethasone was dissolved in AIM V medium at 10 mM and added to the culture at the same time as the allergen at a final concentration of 1 µM.
In preliminary experiments, none of the compounds had a direct effect on the ability of T cells to migrate in response to a number of chemoattractants including IL-8, platelet-activating factor (PAF) and leukotriene B4 (LTB4) (data not shown). When using our protocols for T cell chemotaxis experiments, PAF, IL-8, and LTB4 induce T cell migration in a dose-dependent manner with mean ± SEM chemotactic indices of 6.4 ± 1.2 for 10 nM PAF, 2.3 ± 0.6 for 12.5 nM IL-8 (23), and 5.4 ± 1.8 for 10 nM LTB4 (unpublished observations).
T cell chemotaxis
T cell chemotaxis experiments were performed in 48-well microchemotaxis chambers (Neuro Probe, Cabin John, MD) using T cells from atopic asthmatic donors, none of whom had been treated with corticosteroids. T cells were isolated according to a modification of the protocol described by Stanciu et al. (24). Briefly, PBMC were separated on Lymphoprep, and monocytes were removed by adherence to plastic at 37°C and 5% CO2 for 1 h. After further overnight adherence additional purification of T cells from the remaining nonadherent cells was conducted by negative selection using panning with a mixture of mAb (anti-CD11b, anti-CD14, anti-CD19, and anti-CD16) to remove any contaminating monocytes, B cells, and NK cells, respectively. The cells were finally resuspended in RPMI medium containing 0.4% BSA. T cell purity was >96%, as assessed by flow cytometry and viability was >99%.
Twenty-five microliters of each culture supernatant, or dilutions of human (h-)IL16 and h-RANTES, were placed in the lower wells, which were separated from the corresponding upper wells by an 8-µm pore size polyvinylpyrridone-free polycarbonate membrane. All experiments were performed in triplicate. AIM V medium, supplemented with allergen extract, PSI, CTLA-4-Ig, or dexamethasone, were used as controls. Cell suspensions in 50-µl aliquots containing 2.5 x 105 cells were placed in each upper well, and the chamber was incubated for 60 min at 37°C in 5% CO2. The filter was carefully removed, and its upper side was scraped to remove nonmigrated cells. The migrated cells on the lower side of the membrane were fixed with methanol and stained with May-Gr[umlaut]unwald-Giemsa stain. The number of migrating T cells was counted in five high power fields.
Neutralization experiments
Anti-IL-16 and anti-RANTES Abs were used at 10 and 100 µg/ml, respectively. These were incubated at 37°C for 30 min before chemotaxis with supernatants from Der p-stimulated bronchial biopsies from asthmatic subjects or with hr-IL-16 and hr-RANTES as controls for the specificity of the Abs.
Statistical analysis
All results were corrected for the weight of the mucosal tissue placed in culture by dividing the concentrations of cytokines and the T cell chemotactic activity (expressed as numbers of migrated T cells) by the tissue weight. The spontaneous activity generated by unstimulated tissues was also expressed as an index of spontaneous chemotaxis derived by dividing the number of cells migrating in response to chemoattractants present in supernatants from unstimulated cultures by the numbers of cells migrating spontaneously in response to the corresponding culture medium. The chemotactic activity generated in culture where tissue was stimulated by allergen was also expressed as an index of stimulation of chemotactic activity by Der p (numbers of T cells migrating in response to supernatants from Der p-stimulated cultures divided by the numbers of cells migrating in response to supernatants from unstimulated cultures). Comparisons between treatments with dexamethasone, CTLA-4-Ig, and PSI within the same group were made by the Wilcoxon signed rank test for paired data. Comparisons between asthmatics and control subjects were made using the Mann-Whitney U test for unpaired data. A p value of <0.05 was accepted as statistically significant.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
In 13 of 15 experiments with tissue from asthmatics, the index of spontaneous chemotaxis was >1, indicating the presence of chemotactic activity generated spontaneously by unstimulated tissue (median activity for all experiments, 2.4; range, 1–8.2). This activity was not significantly different from that detected in cultures of tissue from control subjects where an index of >1 was observed in 9 of 12 experiments (median activity for all experiments, 2.7; range, 0.4–6.6).
Both IL-16 and RANTES were detectable in all the experiments, with
considerable variability between subjects (Fig. 1
). The median (range) concentrations of
IL-16, not corrected for tissue weight, were 118.1 pg/ml (18.6–584.9
pg/ml) in asthmatics and 87.7 pg/ml (17.2–139.4 pg/ml) in control
subjects. When corrected for the weight of the biopsies, there was no
significant difference in the median (range) amount of spontaneous
release of IL-16 when comparing cultures of tissue from asthmatics (7.5
pg/mg of tissue (1.1–246.8 pg/mg)) with those from control subjects
(4.8 pg/mg of tissue (2.0–9.4 pg/mg)). Although the concentrations of
spontaneously released RANTES, not corrected for weight, were higher in
the control than in asthmatic subjects (median (range) concentrations
of 188.8 pg/ml (94.5–255.2 pg/ml) and 124.8 pg/ml (2.1–982.0 pg/ml),
respectively), after correcting for tissue weight no significant
difference were noted (in the asthmatics:5.4 pg/mg of tissue
(0.3–124.2 pg/mg), and in control subjects: 15.0 pg/mg of tissue
(6.4–40 pg/mg)).
|
Stimulation of biopsies from asthmatic subjects with Der
p increased the chemotactic activity up to >400%, which was
significantly higher than in cultures of unstimulated tissue from the
same individuals (p = 0.001) (Fig. 1
).
Comparison of median (range) indices of stimulation of chemotactic
activity by Der p showed a significantly
(p = 0.004) higher Der p-generated
activity in the asthmatics (2.7-fold (1.2- to 4.2-fold)) than in
control subjects (1.2-fold (0.2- to 5.1-fold)) (Fig. 1). In 5 of 12
experiments with tissue from control subjects, the indices were <1,
indicating the possible presence of inhibitory activity for T cell
chemotaxis.
Stimulation with Der p induced a significant
(p = 0.04) increase in IL-16 in cultures of
biopsies from asthmatics (to a median (range) of 15.4 pg/mg tissue
(0.8–133.6 pg/mg)) but had no effect on those from control subjects
(median (range), 3.2 pg of tissue (0–22 pg/mg) after stimulation)
(Fig. 1). Comparison of median (range) indices of stimulation of IL-16
released by Der p showed a significant
(p = 0.01) increase in median (range) IL-16
concentrations in cultures of biopsies from asthmatics (2-fold (0.3- to
15.5-fold)) when compared with those from control subjects (1.1 (0- to
3.4-fold)). Der p also induced a significant
(p = 0.02) increase in RANTES release in
culture of biopsies from asthmatics (to a median (range) of 12.9 pg/mg
tissue (0.7–205.3 pg/mg)) without an effect on those from control
subjects (median (range), 11.4 pg/mg tissue (2.5–42.6 pg/mg)) (Fig. 1). The median (range) of indices of stimulation of RANTES produced by
Der p was 1.6-fold (0.3- to 67.8-fold) in the asthmatics and
0.9-fold (0.3- to 1.6-fold) in the control subjects. Comparison of the
magnitude of change after stimulation showed the change in the
asthmatics not to be significantly different from that in control
subjects.
When cytokine generation was expressed as concentrations not corrected for weight of tissue, stimulation of bronchial biopsies from asthmatics induced a median (range) increase from 118.1 pg/ml (18.6–584.9 pg/ml) to 205.2 pg/ml (29.7–497.6 pg/ml) for IL-16 and from 124.8 pg/ml (2.1–982.0 pg/ml) to 157.5 pg/ml (23.6–464.0 pg/ml) for RANTES.
Effect of blocking Abs for IL-16 and RANTES on allergen-induced T cell chemotactic activity
In preliminary experiments with recombinant cytokines, both
hr-IL-16 and hr-RANTES induced T cell migration in a
concentration-dependent manner (Fig. 2),
reaching a plateau at 50 ng/ml, with h-IL-16 being a more potent
chemoattractant. To investigate whether tissue-generated IL-16 and
RANTES accounted for the observed chemotactic activity, supernatants
from cultures of Der p-stimulated tissues were incubated
with excess amounts of neutralizing Abs (10 µg/ml for
anti-IL-16 and 100 µg/ml foranti-RANTES) for 30 min before using
the supernatants in chemotaxis experiments. These concentrations were
shown to block T cell chemotaxis induced by all the concentrations of
recombinant cytokines. Anti-IL-16 and anti-RANTES Abs reduced the
allergen-induced chemotactic activity by a median (range) of 59%
(5–83%) (p = 0.01) and 26% (0–89%)
(p = 0.02), respectively (Fig. 3). The combination of anti-IL-16 and
anti-RANTES Abs caused a significant (p =
0.02) median (range) inhibition of 56% (0–99%), which was not
different from the inhibition achieved with anti-IL-16 Abs
alone.
|
|
Coincubation with dexamethasone of biopsies stimulated with Der p caused a significant inhibition of Der p-induced chemotaxis (median, 48%; range, 0–85%). In addition, dexamethasone inhibited the release of IL-16 in all but two experiments. Median inhibition was 55%, with a maximum of 86%. Similarly, the release of RANTES was inhibited by a median 59%, with a maximum of 85%.
Preincubation of allergen-stimulated bronchial biopsies with the
proteasome inhibitor PSI had a small, albeit significant
(p = 0.03) effect on Der p-induced
generation of T cell chemotactic activity (Fig. 4), with a median 21% inhibition (range,
0–83%). PSI reduced the release of IL-16 by a median of 47% (range,
0–90%) (Fig. 4), but in contrast had no significant effect on
RANTES.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
B in partially
regulating these components of the inflammatory response in asthma. We have demonstrated that the explant model is useful for studying the effects of allergen on bronchial tissue ex vivo, enabling us to begin elucidating the mechanisms regulating the generation of T cell chemoattractants. In maintaining the complex cell-cell interactions, the model not only reflects closely the in vivo situation but also offers several advantages over experimental in vivo challenge in asthmatic volunteers. First, repeat bronchoscopy to sample the airways after allergen challenge is not required. Second, tissue responses of more severe asthmatics, who cannot be challenged with allergen, can be studied. Third, problems of dilution of secreted mediators during BAL are avoided. Fourth, the produced cytokines are not consumed by incoming inflammatory cells. Finally, and most importantly, compounds that cannot be applied in vivo, such as those used in this study, can be used to test concepts of immunoregulation and provide valuable information for drug development.
In the current study, we have focused on IL-16 and RANTES because both
are produced by T cells and may therefore be amenable to inhibition by
agents, such as CTLA-4-Ig, and dexamethasone. We have confirmed
previous observations that allergen induces IL-16 release into the
airways in vivo (8) and that this cytokine is a major
chemoattractant for T cells in asthma. A similar degree of baseline
chemotactic activity was generated spontaneously by tissue from healthy
control and asthmatic subjects, possibly because of the mild disease of
the asthmatics studied. However, stimulation with Der p
increased the chemotactic activity in Der p-sensitized
asthmatics but failed to do the same in nonallergic control subjects,
clearly indicating an allergen-specific effect. A significant part of
this activity was blocked with neutralizing Abs against IL-16 and
RANTES, showing a higher contribution from IL-16 (median, 59%
inhibition) than with RANTES (median, 26% inhibition). The combination
of the two Abs could not completely block the chemotactic activity
induced by stimulation with allergen, indicating the presence of other
chemoattractants. The presence of additional chemoattractants is
further supported by the finding that hr-IL-16 at 560 pg/ml induced a
mean migration of 23 cells/5 high power fields whereas the chemotactic
activity released by allergen-stimulated biopsies induced a mean
migration of 85 cells/5 high power fields and contained only an average
of 214 pg/ml IL-16. One obvious additional chemoattractant is MIP-1,
which we have not been able to assay because of limited amount of
supernatant.
The role of RANTES in allergen-induced T cell recruitment in asthma has been a matter of controversy. Similar levels of protein and mRNA for RANTES have been detected in bronchial biopsies in healthy control and asthmatic subjects (26). This CC chemokine selectively attracts activated/memory CD45RO+ T cells, eosinophils, and monocytes (6) and also induces T cell adherence to adhesion molecules and the extracellular matrix (27). It is of particular interest to asthma pathogenesis because of its preferential recruitment of Th2 cells that bear CCR3. Although RANTES is usually viewed as a late T cell product, our study shows that increased release can occur within 24 h poststimulation with allergen. However, the lack of significance when comparing the magnitude of change between asthmatics and control subjects suggests that it is less important than IL-16. Furthermore, neutralization experiments suggested a relatively minor role for this chemokine when compared with IL-16, at least for the time point (24 h) at which the stimulation was stopped. A role for RANTES bound to proteoglycans within the tissue cannot be excluded, and it is also possible that further extension of culture may reveal additional increases in RANTES. Finally, it is entirely plausible that in vivo recruited cells might cause additional release of RANTES.
The cellular source of the T cell chemotactic activity, IL-16, and RANTES remains to be established. CD4+ and CD8+ T cells, eosinophils, mast cells, and epithelial cells can all generate IL-16. Similarly, RANTES is produced by a variety of cell types including T cells, mast cells, epithelial cells, monocytes, fibroblasts, eosinophils, and platelets. The current study suggests that the generation of these cytokines involves, at least partly, CD28-dependent activation of one or more cells that bear CD28 including T cells, mast cells, and NK cells (20, 21, 28, 29). A role for CD28 has been clearly shown for the process of T cell activation in the context of Ag presentation and interaction with B7 molecules expressed on APC (29). Bioactive IL-16 is produced by CD4+ T cells 18–24 h after stimulation with mitogen, Ag, or anti-CD3 stimulation by a process that requires transcription and translation of new protein (30) and, in view of what is known about the CD28 dependency of T cell responses (17, 28), this is likely to involve costimulation. In that regard, we have recently demonstrated that release of IL-16 from CD4+ T cells is contingent on activation of the ICE enzyme family member, caspase-3 (31, 32). Following TCR stimulation, caspase activation is detected by 16 h, with subsequent release of IL-16 being seen several hours later. Costimulation through CD28 accelerates activation of caspase-3 to detectable levels by 4 h and significantly increases the amount of IL-16 bioactivity beyond that of TCR stimulation alone (32).
Although there is evidence that CD28 may also be involved in the
activation of NK cells (29), little is known about the
mucosal distribution and role of these cells in asthma, and they are
unlikely to be an important cell in allergen-mediated effects. Although
it was reported that IL-16 is stored preformed in bone marrow cultured
human mast cells (33), it is possible that when activated,
mast cells could produce this cytokine either directly or by inducing
its release from resident CD8+ T cells
(9) or epithelial cells (34) via production
of histamine and serotonin (35). Concurrent CD28-dependent
stimulation of murine mast cells and cross-linking of high affinity IgE
receptor (FceRI) greatly enhances TNF- secretion induced via FceRI
(20). Whether human mast cells in bronchial tissue could
use CD28-dependent activation to produce other cytokines/chemokines
such as IL-16 and RANTES needs further verification.
Using PSI, we have shown that at least part (median, 21%) of the
allergen-induced chemotactic response involves NF-B activation. In
accordance with our findings, activation of NF-B via interference
with its cytoplasmic inhibitor I-B has recently been reported to
be induced by the allergen Der p1 in asthmatic bronchial
epithelial cells (36). Our findings suggest, for the first
time, that NF-B could be involved in generating IL-16 by inflamed
tissues, being responsible for at least 50% of the production of this
cytokine. In contrast to a previous report of an NF-B-dependent
expression of RANTES (37), significant inhibition of
RANTES production by PSI could not be achieved, although it is entirely
plausible that higher concentrations of this proteasome inhibitor may
have been effective. In the absence of complete inhibition, a mechanism
not involving the chymotrypsin-like activity of the proteasome cannot
be excluded. The existence of an alternative mechanism would be in
keeping with findings showing that the upstream region of the RANTES
gene contains both AP-1- and NF-B-binding sites (14).
PSI has been shown to inhibit TNF--induced NF-B activation in
HeLa cells, with an IC50 of 15 µM and maximum
inhibition of 75–80% at 100 µM (15). However, more
recent studies on bronchial epithelial cell lines have shown efficacy
already at 5 µM (Dr. John Taylor, Pfizer, unpublished observation).
This suggests that the chymotrypsin-like proteasome activity is
largely, but not exclusively, responsible for NF-B activation.
Because PSI inhibits specifically the chymotrypsin-like activity of the
proteasome, it is possible that NF-B activation was not completely
inhibited in our conditions. Further elucidation of the role of NF-B
would require the use of higher concentrations of PSI, but
concentration/response experiments were not possible in our study
because of limited availability of tissue.
Finally, in keeping with the wide antiinflammatory action of
corticosteroids, dexamethasone caused a stronger inhibition of the T
cell chemoattractant activity generated by allergen (48%) than the
inhibition with PSI (21%). Possible mechanisms involved in this
inhibition include binding of NF-B by the glucocorticoid
receptor/glucocorticoid complex and increased stimulation of IB
synthesis, both of which prevent NF-B from migrating into the
nucleus (38) and also inhibition of activation of AP-1
(39). Whereas expression of the gene for RANTES is
dependent on activation of both NF-B and AP-1 (14), the
transcription factors regulating IL-16 synthesis are unknown.
Nevertheless, our observations are in agreement with a recent study
showing that corticosteroids inhibit IL-16 expression during the
allergen-induced late phase nasal response in allergic rhinitics
(40). Similarly, our findings are in accordance with the
inhibitory effects both in vitro and in vivo of the topical
corticosteroid beclomethasone dipropionate on the expression of RANTES
by bronchial epithelial cells (41).
In conclusion, we have demonstrated that allergen causes up-regulation
of T cell chemoattractant activity in the airways of asthmatic
individuals which can be inhibited by corticosteroids and, to a lesser
extent, inactivation of NF-B. A proportion of the response involving
both IL-16 and RANTES requires cell activation via the costimulatory
molecule CD28.
|
Acknowledgments |
|---|
|
Footnotes |
|---|
2 Address correspondence and reprint requests to Dr. R. Djukanovi, Respiratory Cell and Molecular Biology Division, University Medicine, Level D, Centre Block, Southampton General Hospital, Tremona Road, Southampton SO16 6YD, U.K. E-mail address:
3 Abbreviations used in this paper: BAL, bronchoalveolar lavage; MIP, macrophage-inflammatory protein; PSI, Cbz-Ile-Glu(OtBu)-Ala-leucinal; PAF, platelet-activating factor; LTB4, leukotriene B; hr, human recombinant; Fc
RI, high affinity IgE receptor.
Received for publication July 16, 1999. Accepted for publication October 13, 1999.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
, R., I. Feather, C. Gratziou, A. F. Walls, D. Peroni, P. Bradding, M. Judd, P. H. Howarth, S. T. Holgate. 1996. The effect of natural allergen exposure during the grass-pollen season on airways inflammatory cells and asthma symptoms. Thorax 51:575. (MIP1) in bronchoalveolar lavage fluid of antigen-challenged asthmatics. Am. J. Respir. Cell Mol. Biol. 13:738.[Abstract]
B and stabilizes a newly phosphorylated form of IB- that is still bound to NF-B. EMBO J. 13:5433.[Medline]
, S. T. Holgate. 1997. B7 costimulation is required for IL-5 and IL-13 secretion by bronchial biopsy tissue of atopic asthmatic subjects in response to allergen stimulation. Am. J. Respir. Cell Mol. Biol. 20:153. by costimulation of mast cells via CD28 and FceRI. J. Immunol. 158:2382.[Abstract]
, R., J. W. Wilson, C. K. W. Lai, S. T. Holgate, P. H. Howarth. 1991. The safety aspects of fiberoptic bronchoscopy, bronchoalveolar lavage, and endobronchial biopsy in asthma. Am. Rev. Respir. Dis. 143:772.[Medline]
. Phosphodiesterase and cyclic AMP-dependent inhibition
of T lymphocyte chemotaxis. Eur. Respir. J. In press.
. 1996. Production of IL-8 and IL-4 by positively and negatively selected CD4+ and CD8+ human T cells following a four-step cell separation method including magnetic cell sorting (MACS). J. Immunol. Methods 189:107.[Medline]
, R., S. Homeyard, C. Gratziou, J. Madden, A. Walls, S. Montefort, D. Peroni, R. Polosa, S. T. Holgate, P. H. Howarth. 1997. The effect of treatment with oral corticosteroids on asthma symptoms and airway inflammation. Am. J. Repsir. Crit. Care Med. 155:826.[Abstract]
B activation through interference with IB function in asthmatic bronchial epithelial cells. Biochem. Biophys. Res. Commun. 236:522.[Medline]
B potently up-regulates the promoter activity of RANTES, a chemokine that blocks HIV infection. J. Immunol. 158:3483.[Abstract]
B activity through induction of IB synthesis. Science 270:286.This article has been cited by other articles:
|
|
 |
|
  T. Kallinich, K. C. Beier, U. Wahn, P. Stock, and E. Hamelmann T-cell co-stimulatory molecules: their role in allergic immune reactions Eur. Respir. J., June 1, 2007; 29(6): 1246 - 1255. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Poster presentations Thorax, December 1, 2006; 61(suppl_2): ii57 - ii133. [Full Text] [PDF]
|
|
|
|
 |
|
 A. Nencioni, K. Schwarzenberg, K. M. Brauer, S. M. Schmidt, A. Ballestrero, F. Grunebach, and P. Brossart Proteasome inhibitor bortezomib modulates TLR4-induced dendritic cell activation Blood, July 15, 2006; 108(2): 551 - 558. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. K. Wong, L. C. W. Lit, L. S. Tam, E. K. Li, and C. W. K. Lam Aberrant production of soluble costimulatory molecules CTLA-4, CD28, CD80 and CD86 in patients with systemic lupus erythematosus Rheumatology, August 1, 2005; 44(8): 989 - 994. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. F. Little, E. Lynch, G. Fine, D. M. Center, and W. W. Cruikshank Tumor Necrosis Factor-{alpha}-Induced Synthesis of Interleukin-16 in Airway Epithelial Cells: Priming for Serotonin Stimulation Am. J. Respir. Cell Mol. Biol., March 1, 2003; 28(3): 354 - 362. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 O. Fahy, S. Senechal, J. Pene, A. Scherpereel, P. Lassalle, A.-B. Tonnel, H. Yssel, B. Wallaert, and A. Tsicopoulos Diesel Exposure Favors Th2 Cell Recruitment by Mononuclear Cells and Alveolar Macrophages from Allergic Patients by Differentially Regulating Macrophage-Derived Chemokine and IFN-{gamma}-Induced Protein-10 Production J. Immunol., June 1, 2002; 168(11): 5912 - 5919. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. F. Little, W. W. Cruikshank, and D. M. Center IL-9 Stimulates Release of Chemotactic Factors from Human Bronchial Epithelial Cells Am. J. Respir. Cell Mol. Biol., September 1, 2001; 25(3): 347 - 352. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. S. Burr, S. L. Kimzey, D. R. Randolph, and J. M. Green CD28 and CTLA4 Coordinately Regulate Airway Inflammatory Cell Recruitment and T-Helper Cell Differentiation after Inhaled Allergen Am. J. Respir. Cell Mol. Biol., May 1, 2001; 24(5): 563 - 568. [Abstract] [Full Text]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
) subjects following 24-h stimulation with
Der p (2000 U/ml) or cultured in medium alone (M) as
control. The horizontal lines denote medians for all experiments. The
experiments were performed as described in Materials and
Methods.
